Antibody glycoengineering for drug delivery applications
2019-12
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Antibody glycoengineering for drug delivery applications
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2019-12
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Monoclonal antibodies (mAbs) are frontline drugs for the treatment of many diseases including cancer 1 and rheumatoid arthritis 2 . In addition to their natural role as neutralizers of pathogens and toxins as well as in the recruitment of immune elements (complement, improving phagocytosis, antibody dependent cytotoxicity), they can be used as carriers for tumor-targeted delivery of therapeutic and diagnostic agents 3 . However, conjugation of drug or drug-encapsulated nanoparticles to antibodies can often result in reduced affinity of the antibody towards the target antigen. The overall objective of this thesis is to advance a new antibody glycoengineering technology that will allow for facile synthesis of antibody-based drug delivery systems. Most therapeutic mAbs are of the IgG class, which contains a glycosylation site in the Fc region at position 297 4 . In chapter 2, we investigated a glycoengineering strategy that enables the introduction of artificial azide groups at this glycosylation site without affecting their antigen affinity. This is based on the observation that glycosyltransferases present in mammalian cells can incorporate non-natural sugars (e.g., azido mannose) at glycosylation sites on an IgG molecule during the post translational modification. The azide groups in these artificial sugars are then available to react with alkynes through copper-catalyzed ‘click’ chemistry or with strained alkynes such as dibenzyl cyclooctyne (DBCO) allowing for biorthogonal, copper-free ‘click’ chemistry. Because the sugars are added reproducibly and at a site that does not affect antigen binding, the glycoengineering technology would overcome problems associated with traditional conjugation strategies. Using this approach, azide groups were introduced in anti-CD133 and anti-perlecan (AM6) antibodies. Further, the azide groups were available to react with various DBCO conjugates including fluorophores, drug molecules and nanoparticles. Importantly, the addition of artificial sugar and subsequent azide-alkyne reaction did not affect the affinity of the antibody for the target antigen. Antibody–drug conjugates (ADCs) have emerged as the next generation anticancer therapeutic agents. In chapter 3, glycoengineered AM6 antibody was used to generate an ADC with monomethyl auristatin E (MMAE) as the cytotoxic drug. The glycoengineering approach resulted in an ADC with a DAR of 2-3 drug molecules per antibody. The AM6- MMAE conjugate demonstrated enhanced cell kill in vitro and significantly improved anticancer efficacy in vivo compared to free MMAE. Similarly, in chapter 4, glycoengineered AM6 antibody was used to generate antibody conjugated polymeric nanoparticles loaded with paclitaxel. These perlecantargeted nanoparticles showed enhanced antitumor efficacy in vitro and in vivo in TNBC tumor models. Similarly, antibody conjugated nanoparticles showed enhanced antitumor efficacy in vitro and complete tumor growth inhibition in vivo in a non-muscle invasive bladder cancer model. We expect that this glycoengineering strategy will prove to be a unique platform technology that will have a significant impact on antibody-based therapeutics.
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University of Minnesota Ph.D. dissertation. December 2019. Major: Pharmaceutics. Advisor: Jayanth Panyam. 1 computer file (PDF); xiv, 110 pages.
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Sehgal, Drishti. (2019). Antibody glycoengineering for drug delivery applications. Retrieved from the University Digital Conservancy, https://hdl.handle.net/11299/223122.
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